HMGB proteins are members of the High Mobility Group (HMG) superfamily, possessing a unique DNA-binding domain, the HMG-box, which can bind non-B-type DNA structures (bent, kinked and unwound) with high affinity, and also distort DNA by bending/looping and unwinding. HMGBs (there are four HMGBs in mammals, HMGB1-4) are highly abundant and ubiquitously expressed non-histone proteins, acting as DNA chaperones influencing multiple processes in chromatin such as transcription, replication, recombination, DNA repair and genomic stability. Although HMGB1 is a nuclear protein, it can be secreted into the extracellular milieu as a signaling molecule when cells are under stress, in particular, when necrosis occurs. Mammalian HMGBs contain two HMG-boxes arranged in tandem, share more than 80% identity and differ in the length (HMGB1-3) or absence (HMGB4) of the acidic C-tails. The acidic tails consist of consecutive runs of only Glu/Asp residues of various length, and modulate the DNA-binding properties and functioning of HMGBs. HMGBs are subject to post-translational modifications which can fine-tune interactions of the proteins with DNA/chromatin and determine their relocation from the nucleus to the cytoplasm and secretion. Association of HMGBs with chromatin is highly dynamic, and the proteins affect the chromatin fiber as architectural factors by transient interactions with nucleosomes, displacement of histone H1, and facilitation of nucleosome remodeling and accessibility of the nucleosomal DNA to transcription factors or other sequence-specific proteins.